CN104542256A - Breeding method for importing false smut resistance of indica rice into japonica rice - Google Patents
Breeding method for importing false smut resistance of indica rice into japonica rice Download PDFInfo
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Abstract
The invention discloses a breeding method for importing false smut resistance of indica rice into japonica rice. The method includes the following steps: (1) selecting japonica rice varieties/series to be improved and false smut resistant resources of the indica rice; (2) identifying the crude toxin tolerance concentration of the selected japonica rice varieties/series to be improved; (3) cross-breeding to obtain three-way cross coeno-species F1; (4) culturing anther of the three-way cross coeno-species F1 in a culture medium for culturing rows of indica rice; (5) field-planting flower cultured seedlings: planting a single anther cultured seedling to obtain a plurality of japonica rice flower cultured families with the false smut resistance improved, and further planting different flower cultured families in the field and screening to obtain new japonica rice varieties with good agronomic shape and improved false smut resistance, using the japonica rice varieties/series to be improved as a contrast. The breeding method provided by the invention can effectively import the false smut resistance of indica rice into japonica rice, and the false smut resistance breeding can be finished in about three years, therefore, the normal false smut resistance breeding period is greatly shortened; the breeding method has a great breeding application value.
Description
Technical field
The present invention relates to a kind of Japonica two line hybrid rice method, refer in particular to a kind of by special for long-grained nonglutinous rice false smut resistance and import the breeding method of japonica rice efficiently, belong to technical field of agriculture science.
Background technology
Rice green smut (rice false smut), having another name called pseudo-smut, green smut, paddy blossom disease, blue or green powder disease and good harvest disease etc., is one of important disease in Rice Production.False smut occurs the most frequent in Rice In Asian Cultivated Rice district, as countries in Southeast Asia such as China, India, Burma, also all have generation in areas such as Europe, America and Africa.False smut is not serious to the damage losses of paddy rice in history, as just the Minor diseases of Rice Production.But in recent years, along with the raising of Global climate change and China's high yield and high quality hybrid rice spread and paddy rice liquid manure level, false smut is in the large generation in multiple province such as Hunan, Jiangxi, Liaoning, Yunnan, Heilungkiang that is Japanese and China, present the features such as morbidity scope is wide, frequency of disease development is high, production loss is serious, harm increases the weight of year by year.False smut rises to Major Diseases from Minor diseases, becomes the important disease of the another serious threat paddy rice high-quality after rice blast, stripe virus disease, high yield.
After paddy rice generation false smut, a large amount of flat granulosis grain can be caused, reduce rice yield, the general underproduction 10% ~ 20%, and containing a large amount of coloured toxin in rice curve, these toxin not only significantly can reduce rice quality, and very harmful to health.Research finds, the tubulin of this toxin to animal has obvious inhibitory action, feeds mouse, can cause the necrosis such as rat liver, kidney and bladder, and can cause people and animals' chronic poisoning with the paddy containing rice false smut.Rice green smut have impact on China's grain stable high yield and grain and food safety greatly.The control of false smut and disease-resistant research become the problem paid close attention at present and urgently research and solve gradually.
For a long time, chemical agent and cultivation step are depended on to the control of rice green smut, but control efficiency is not satisfactory.Integrated control technique is taked to prevent the generation of false smut in present agricultural production, Main Means takes some preventive measures, antibiotic and sterilizing means and applying pesticides is adopted to carry out pharmacotherapy as selected in disease-resistant variety, cultivation, and set up false smut forecast system, long-term production practices prove, the Breeding and application of rice green smut resistant variety prevents and treats the most economical effective measures of false smut.
Indica rice breaks up and significant difference deeply, and make them differ from one another in output, quality and resistance etc., no matter its Hybrids F1, all have powerful hybrid vigour at nutrition organs or on yield traits.Yuan Longping proposes the imagination of subspecies indica and japonica hybrid use of advantage first, think and directly utilize inter-subspecific heterosis to be to be hopeful most during output breaks through and can compared with the approach obtaining effect in short-term, propose application wide compatibility gene and phto-temperature reaction and realize the strategy flexibility that inter-subspecific heterosis directly utilizes.Rice Cross Breeding Between Indica And Japonica will break the boundary of breeding in subspecies, and in subspecies, subspecies breeding is moved towards in breeding.Indica-japonica hybrid is sub-distant hybridization, and filial generation easily produces huge variation and heterobeltiosis, contains huge genetic potential.Indica-japonica hybridization conventional breeding and subspecies indica and japonica hybrid use of advantage have become important means and the method for the research of current Rice Super-yield Breeding.The breeding practice of China also proves, utilizes Indica-japonica hybridization to be create the effective method of novel strain type excellent germplasm.Current Xian round-grained rice hands over the kind be bred as to have the short round-grained rice 23 in Zhejiang, T209 and city spy 232 etc., No. 5, the distant round-grained rice in Liaoning, distant round-grained rice 326, Shen Nong 1033, and the Zijin round-grained rice in Jiangsu, makes system in Pekinese, and Hubei Province in Hubei educates late No. 5 etc.These kinds (being) show the good characteristic of high yield, disease-resistant and high-quality, but some Hybrids of Indica and Japonica Posterity phenotypes go out some shortcomings, as the not blast resisting of " thousandweight wave ", " southern round-grained rice 35 " be easy to Spike sprouting etc.
Abroad indica-japonica hybrid traditional breeding method is also paid close attention.From the middle and later periods sixties, the indica-japonica hybrid conventional breeding of Korea S achieves remarkable achievement, and be bred as the high yield Dwarf-lines systems such as unification, water are former, close sun, these kinds are generally than traditional round-grained rice type kind volume increase 20% ~ 40%.Japan introduces the Indica Rice kind on the ground such as China and Korea S, hybridize with local japonica rice variety, increase the floret bears of hybrid generation, improve resistance and the yield stability of filial generation simultaneously, successively 52 new lines likely and 5 new varieties are bred as, as China 91, northern land 125, northern land 142, the Northeast 146, Xing Feng, Qiu Li etc., than check variety volume increase 20% ~ 36%.Breeding man of Japan is thought, the index that will realize volume increase 50% in the phase III must rely on hybrid rice, and particularly after paddy rice wide compatibility gene theory proposes, the main subspecies indica and japonica hybrid use of advantage approach that relies on reaches.
But, because Xian, round-grained rice belong to two subspecies, although Fl nutritional advantages is very strong.But generally all there is part reproductive disorder, the plant height of subspecies indica and japonica hybrid F1 generation is higher, and breeding time is super close late-maturing, and the abortion of pollen part, the problem such as ripening rate is on the low side, be the Three Obstacles that Xian round-grained rice friendship breeding faces, seriously limit the direct utilization of this advantage on producing.No matter Xian round-grained rice is conventional rice variety seed selection or Breeding for restoration lines if being handed over and utilized, be substantially to be based upon on random combo and a large amount of basis selected, and needs to expend years and the huge amount of labour.
Research finds, typical Xian Xian hands over and round-grained rice round-grained rice is transferred in gene exchange few, and Fl advantage is weak, and excessive Xian round-grained rice gene exchange is also unfavorable for that F1 is nourishing and growing and the coordination on reproductive growth, be difficult to equally show yield heterosis, this just relates to the degree problem of Xian round-grained rice gene introgression.The breeding fact shows, Xian round-grained rice gene introgression has played important function in Breeding on Super Hybrid Rice, and by Molecular Detection, the parent of a lot of super hybridized rice is all obtained by Xian round-grained rice gene introgression.But gene introgression in Super rice breeding, or is realized by the hybridization in repeatedly generation, the screening that backcrosses, and breeding cycle is long, and workload is huge.
Indica rice breaks up and significant difference deeply, them are made to differ from one another in output, quality and resistance etc., one of them is exactly the Resistant expression difference highly significant false smut germ, rice variety will far away higher than japonica rice variety to the resistance level of false smut, and japonica rice shows to lack in japonica rice genome ubiquitous anti-false smut gene in long-grained nonglutinous rice genome to the resistance level that false smut is poor.How the resistant gene in high resistance false smut long-grained nonglutinous rice can be shifted by accurate means efficiently and to cultivate anti-false smut japonica rice new varieties into japonica rice genome, production has high value.
Along with the development of biotechnology, many new breeding techniques have been there is again except conventional cross-breeding, this give Rice Resistance false smut breeding new way, one utilizes false smut culture fluid to extract Raw toxin as Selective Pressure, carry out screening mutant on a cellular level, and be combined with traditional breeding method and cultivate new disease-resistant varieties, but low by the frequency of Raw toxin induced mutations, screening amount is large; The molecular breeding technology that the second is exactly is representative with transgenic technology and molecular marker assisted selection breeding technique.The appearance of molecular breeding technology, breeding difficulty is handed over to be a kind of feasible solution originally to the breeding of Rice Resistance false smut and Xian round-grained rice, at present, transgenic breeding and molecular marker assisted selection equimolecular technology Breeding are the new roads of current agricultural development, are known as the appearance of the third time agriculture green revolution by scientific circles.But, specific excellent main breed is carried out to the transgenic breeding of anti-false smut gene and other excellent genes, originally accurately can realize the improvement that Xian round-grained rice is handed over, but be resisted due to current transgenic breeding, and first molecular marker assisted selection breeding needs gene location, the hybridization of land for growing field crops many generations is needed to backcross equally, the problem that Xian round-grained rice is handed over can not be solved, in addition, molecular engineering breeding, all exist costly, problem that technical threshold is high, to be difficult to grasp by general breeder.
Summary of the invention
The Hybrids F1 plant height that the object of the invention is to hand over conventional breeding to face for existing Xian round-grained rice is higher, breeding time super close late-maturing, obstacle that pollen part abortion ripening rate is on the low side, provide a kind of by special for long-grained nonglutinous rice false smut resistance and import japonica rice efficiently thus improve the breeding method of japonica rice variety.
The object of the invention is to solve by the following technical programs:
Long-grained nonglutinous rice false smut resistance efficiently imports a breeding method for japonica rice, it is characterized in that: the step of this breeding method is as follows:
(1) choose and wait to improve japonica rice variety/be and the anti-anti-source of false smut long-grained nonglutinous rice: choose the good japonica rice variety of economical character/be and to the japonica rice variety chosen/be the Fiber differentiation carrying out arranging Xian medium, final selected induction of anther callus rate higher than 25% japonica rice variety/be as treating improvement japonica rice variety/be, choose the good and rice variety containing false smut high resistance gene of economical character/be as the anti-anti-source of false smut long-grained nonglutinous rice;
(2) improvement japonica rice variety/be the qualification carrying out Raw toxin patience concentration is treated to what choose: the Extracted toxin liquid adding certain gradient concentration respectively in differential medium, differentiation cultivation is carried out to the anther callus waiting to improve japonica rice variety/be selected in step (1), and selects the Extracted toxin liquid of suitable screening pressure level concentration as japonica rice anther culture Raw toxin Selective Pressure;
(3) crossbreeding obtains three and hands over F1 crossbreed: the anti-anti-source of false smut long-grained nonglutinous rice of screen field planting and Wide compability material Xian round-grained rice are built bridge and obtained Hybrid F1, the flower pesticide choosing Hybrid F1 again with wait to improve japonica rice variety/be and hybridize and obtain crossbreed three and hand over F1;
(4) F1 crossbreed is handed over to carry out row Xian medium anther culture to three: three hand over the positive season sowing of F1 crossbreed, choose healthy and strong main Honoka medicine and carry out row Xian medium flower pesticide Fiber differentiation, obtain anther callus, and the anther callus selecting diameter 0.4 ~ 0.8cm is inoculated in the differential medium that with the addition of Selective Pressure concentration Raw toxin, temperature 27 ~ 29 DEG C, cultivate under the condition of the dark 10h of light 14h/, obtain flower training seedling, treat that bloom control is transferred in root media and carried out squamous subculture after growing to 2cm, growing to until bloom control to transplant to Turnover Box strong sprout after 8cm, land for growing field crops is transplanted into after 10 ~ 15 days,
(5) bloom control individual plant sowing obtains flower training family, further economical character, false smut resistance screening obtain the Keng rice lines of improvement: field planting bloom control, the japonica rice training family that individual plant sowing obtains the improvement of false smut resistance is some, the further field seeding of training family of not suiting, to treat that improvement japonica rice variety/be parent is for contrast, screening obtains the Keng rice lines that the good and false smut resistance of agronomic shape obtains improveing.
The japonica rice variety that economical character in described step (1) is good/be refers to that output is high, good grain quality, false smut resistance need to be improved and plant height should not too high, breeding time is precocious or in ripe japonica rice variety/be; The anti-anti-source of false smut long-grained nonglutinous rice refers to that output is high, the rice variety of good grain quality, high resistance false smut/be.
The process that the japonica rice variety that the economical character chosen in described step (1) is good/be carries out the Fiber differentiation arranging Xian medium is as follows:
(1.1) to draw materials and pretreatment: by japonica rice variety good for the economical character chosen/be land for growing field crops conventional cultivation, get pulvinus apart from be 5 ~ 7cm, fringe look be pistac, flower pesticide is faint yellow and length be about clever shell 1/3 ~ 1/2, the main fringe of stalwartness that do not split of booting is for for trying material, retain 2 ~ 3 blades, behind 75% alcohol wipe surface, wrap up and overcoat plastic sack with wet towel, Cold pretreatment 3 days at putting 4 DEG C;
(1.2) inoculate: after pretreatment, rice bud containing main fringe is cut, then this rice bud is soaked in the alcohol of 75% and processes 3 ~ 4min, then this rice bud is shifted and peel off main fringe in superclean bench, strip uniform ramuscule stalk sized by small ear further, the following rear concentration of gauze parcel ramuscule stalk of sterilizing is the mercuric chloride sterilizing 10min of 0.1%, rinsed with sterile water 3 ~ 4 times, open gauze to dry, finally cut off flower pesticide top with the scissors of sterilizing, tremble medicine method inoculation flower pesticide in row Xian medium;
(1.3) Fiber differentiation: dark surrounds evoked callus row Xian medium being placed in 24 ~ 26 DEG C, white-flaxen unformed cell mass is there is at the edge of flower pesticide after 30 ~ 40 days, and add up induction of anther callus rate afterwards flower pesticide inoculated and cultured 50 days, selected induction of anther callus rate higher than the good japonica rice variety of the economical character of 25%/be as treating improvement japonica rice variety/be.
Rice bud in described step (1.2) refers in the region of rice bud top pillow distance for internode top 0.5cm under 7cm to fringe the part of cutting material and obtaining, and the upper flower pesticide quantity of the ramuscule stripped stalk is 15 ~ 25.
Row Xian culture medium prescription in described step (1.2) is: N6 macroelement+N6 trace element+N6 organic matter+2mg/L2,4-D+0.5mg/LNAA+0.5mg/L protein hydrolysate+5% sucrose+0.8% plant gel, and the pH value of row Xian medium is 5.8.
Raw toxin patience concentration qualification process in described step (2) is as follows:
(2.1) preparation of Raw toxin: mixing rice false smut is inoculated in plane expand breeding culture medium expands numerous, be transferred to after growing mycelium and produce spore expand in PDA liquid nutrient medium numerous, 28 DEG C of lucifuge shaken cultivation use sterile gauze elimination bacterium after 20 days, culturing filtrate is after the centrifugal 15min of 3000rpm, get supernatant and make crude extract at 121 DEG C of high pressure moist heat sterilization 20min, the bacteria-free filtrate that obtains is rice false smut Raw toxin;
(2.2) treat that improvement japonica rice variety/be carries out the qualification of Raw toxin endurance concentration to what choose: the Extracted toxin liquid adding certain gradient concentration respectively in differential medium, then the anther callus waiting to improve japonica rice variety/be Fiber differentiation selected in step (1) is shifted into differential medium, Fiber differentiation added up phenylacetic acid after 30 ~ 40 days, and selected the Extracted toxin liquid of suitable screening pressure level concentration to hand over F1 crossbreed anther culture Raw toxin Selective Pressure concentration as three.
Suitable screening pressure level concentration in described step (2.2) refers to the selected Raw toxin concentration of anther callus when being 0.8% ~ 1.2% containing the differentiation rate in the differential medium of Raw toxin until improvement japonica rice variety/be Fiber differentiation.
Plane in described step (2.1) expands breeding culture medium formula: yeast extract 2.5g/L+ soluble starch 10g/L+ agar powder 16g/L, and pH value is 6.0; PDA liquid culture based formulas is potato 200g/L+ glucose 20g/L+ agar 17g/L, and pH value is 6.5; Differential medium formula in described step (2.2) is: MS minimal medium+2mg/L6-BA+0.2mg/LNAA+ gradient concentration Extracted toxin liquid, the pH value of differential medium is 6.0.
In described step (4) three hand over the induction of anther callus rate of F1 crossbreed lower than 3% time, F1 crossbreed and japonica rice parent need be handed over to backcross by three and obtain BC1F1 crossbreed, the positive season sowing of BC1F1 crossbreed, choose healthy and strong main fringe and re-start row Xian medium flower pesticide Fiber differentiation, until obtain the anther callus that inductivity is not less than 3%, the anther callus selecting diameter 0.4 ~ 0.8cm is inoculated in the differential medium adding Raw toxin Selective Pressure, temperature 27 ~ 29 DEG C, cultivate under the condition of the dark 10h of light 14h/, obtain flower training seedling, treat that bloom control is transferred in root media and carried out squamous subculture after growing to 2cm, growing to until bloom control to transplant to Turnover Box strong sprout after 8cm, land for growing field crops is transplanted into after 10 ~ 15 days.
Row Xian culture medium prescription in described step (4) is: N6 macroelement+N6 trace element+N6 organic matter+2mg/L2,4-D+0.5mg/LNAA+0.5mg/L protein hydrolysate+5% sucrose+0.8% plant gel, and the pH value of row Xian medium is 5.8; Differential medium formula is: the Extracted toxin liquid of MS mineral salt+MS organic matter+suitable screening pressure level concentration of sucrose 3%+ agar 0.7%+KT2.0mg/L+NAA0.5mg/L+ protein hydrolysate 1g/L+, and the pH value of differential medium is 5.8; Prescription of rooting medium is: a large amount of+Ms trace+Ms organic matter+sucrose 3%+ agar 0.8% of 1/2Ms, the pH value of root media is 5.8.
The present invention has the following advantages compared to existing technology:
Breeding method of the present invention, compared with conventional breeding methods, by specific for long-grained nonglutinous rice false smut resistance and import japonica rice efficiently, can greatly reduce screening operation amount, within about 3 years, can complete breeding, significantly shorten conventional anti-false smut breeding cycle; Compared with molecular engineering breeding, there is again the advantage that operation threshold is relatively low, spend less and breeding cycle to shorten, there is larger Breeding Application and be worth.
Accompanying drawing explanation
Accompanying drawing 1 is breeding method Technology Roadmap of the present invention.
Embodiment
Below in conjunction with accompanying drawing 1, the present invention is further illustrated with embodiment.
As shown in the Technology Roadmap of Fig. 1: a kind of long-grained nonglutinous rice false smut resistance efficiently imports the breeding method of japonica rice, and the step of this breeding method is as follows:
(1) choose and treat improvement japonica rice variety/be and the anti-anti-source of false smut long-grained nonglutinous rice:
Choose the good japonica rice variety of economical character/be, because Xian round-grained rice is handed over, F-1 hybrids has that plant height is higher usually, breeding time super close late-maturing feature, therefore the good japonica rice variety of this economical character/be refers to that output is high, good grain quality, false smut resistance need improvement and plant height should not too high, breeding time is precocious or in ripe japonica rice variety/be, then the Fiber differentiation arranging Xian medium is carried out to the japonica rice variety chosen/be, whether flower pesticide evoked callus is carried out in the row of being applicable to Xian medium to check this japonica rice variety/be, the process of Fiber differentiation is as follows: (1.1) draw materials and pretreatment: by japonica rice variety good for the economical character chosen/be land for growing field crops conventional cultivation, getting pulvinus distance is 5 ~ 7cm, fringe look is pistac, flower pesticide is faint yellow and length is about 1/3 ~ 1/2 of clever shell, the main fringe of stalwartness that booting is not split is for supplying examination material, retain 2 ~ 3 blades, behind 75% alcohol wipe surface, wrap up and overcoat plastic sack with wet towel, Cold pretreatment 3 days at putting 4 DEG C, (1.2) inoculate: after pretreatment, rice bud containing main fringe is cut, rice bud refers in the region of rice bud top pillow distance for internode top 0.5cm under 7cm to fringe the part of cutting material and obtaining, then this rice bud is soaked in the alcohol of 75% and processes 3 ~ 4min, then this rice bud is shifted and peel off main fringe in superclean bench, strip uniform ramuscule stalk sized by small ear further, the upper flower pesticide restricted number of each ramuscule stalk is 15 ~ 25, following sterilized gauze parcel ramuscule stalk rear concentration be 0.1% mercuric chloride sterilizing be not less than 10min, rinsed with sterile water 3 ~ 4 times, open gauze to dry, finally cut off flower pesticide top with the scissors of sterilizing, tremble medicine method inoculation flower pesticide in row Xian medium, row Xian culture medium prescription is: N6 macroelement+N6 trace element+N6 organic matter+2mg/L2, 4-D+0.5mg/LNAA+0.5mg/L protein hydrolysate+5% sucrose+0.8% plant gel, the pH value of this row Xian medium is 5.8, (1.3) Fiber differentiation: dark surrounds evoked callus row Xian medium being placed in 24 ~ 26 DEG C, white-flaxen unformed cell mass is there is at the edge of flower pesticide after 30 ~ 40 days, and add up induction of anther callus rate afterwards flower pesticide inoculated and cultured 50 days, selected induction of anther callus rate higher than the good japonica rice variety of the economical character of 25%/be as treating improvement japonica rice variety/be.Choose the good and rice variety containing false smut high resistance gene of economical character/be as the anti-anti-source of false smut long-grained nonglutinous rice simultaneously, the anti-anti-source of false smut long-grained nonglutinous rice refers to that output is high, the rice variety of good grain quality, high resistance false smut/be, guarantees that false smut high resistance gene is contained in the anti-source of this long-grained nonglutinous rice.
(2) improvement japonica rice variety/be the qualification carrying out Raw toxin patience concentration is treated to what choose:
Raw toxin patience concentration qualification process is as follows: the preparation of (2.1) Raw toxin: mixing rice false smut is inoculated in plane expand breeding culture medium expands numerous, plane expands breeding culture medium formula: yeast extract 2.5g/L+ soluble starch 10g/L+ agar powder 16g/L and pH value is 6.0, be transferred to after growing mycelium and produce spore expand in PDA liquid nutrient medium numerous, PDA liquid culture based formulas is potato 200g/L+ glucose 20g/L+ agar 17g/L, pH value is 6.5, 28 DEG C of lucifuge shaken cultivation use sterile gauze elimination bacterium after 20 days, culturing filtrate is after the centrifugal 15min of 3000rpm, get supernatant and make crude extract at 121 DEG C of high pressure moist heat sterilization 20min, the bacteria-free filtrate that obtains is rice false smut Raw toxin, (2.2) treat that improvement japonica rice variety/be carries out the qualification of Raw toxin endurance concentration to what choose: the Extracted toxin liquid adding certain gradient concentration respectively in differential medium, then the anther callus waiting to improve japonica rice variety/be Fiber differentiation selected in step (1) is shifted into differential medium, differential medium formula is: MS minimal medium+2mg/L6-BA+0.2mg/LNAA+ gradient concentration Extracted toxin liquid and the pH value of differential medium is 6.0, Fiber differentiation added up phenylacetic acid after 30 ~ 40 days, and select the Extracted toxin liquid reaching suitable screening pressure level concentration to hand over F1 crossbreed anther culture Raw toxin Selective Pressure concentration as three, wherein suitable screening pressure level concentration refers to the selected Raw toxin concentration of anther callus when being 0.8% ~ 1.2% containing the differentiation rate in the differential medium of Raw toxin until improvement japonica rice variety/be Fiber differentiation.
(3) crossbreeding obtains three and hands over F1 crossbreed:
The anti-anti-source of false smut long-grained nonglutinous rice of screen field planting and Wide compability material Xian round-grained rice are built bridge and are obtained Hybrid F1, the flower pesticide choosing Hybrid F1 again with wait to improve japonica rice variety/be and hybridize and obtain crossbreed three and hand over F1.
(4) F1 crossbreed is handed over to carry out row Xian medium anther culture to three:
Three hand over the positive season sowing of F1 crossbreed, choose healthy and strong main Honoka medicine and carry out row Xian medium flower pesticide Fiber differentiation acquisition anther callus, row Xian culture medium prescription is: N6 macroelement+N6 trace element+N6 organic matter+2mg/L2,4-D+0.5mg/LNAA+0.5mg/L protein hydrolysate+5% sucrose+0.8% plant gel, the pH value of row Xian medium is 5.8, when three hand over the induction of anther callus rate of F1 crossbreed lower than 3% time, F1 crossbreed and japonica rice parent need be handed over to backcross by three and obtain BC1F1 crossbreed, the positive season sowing of BC1F1 crossbreed, choose healthy and strong main fringe and re-start row Xian medium flower pesticide Fiber differentiation, until obtain the anther callus that inductivity is not less than 3%, the anther callus selecting diameter 0.4 ~ 0.8cm is inoculated in the differential medium that with the addition of Selective Pressure concentration Raw toxin, differential medium formula is: the Extracted toxin liquid of the gradient concentration of MS mineral salt+MS organic matter+sucrose 3%+ agar 0.7%+KT2.0mg/L+NAA0.5mg/L+ protein hydrolysate 1g/L+ screening pressure level and the pH value of differential medium is 5.8, temperature 27 ~ 29 DEG C, cultivate under the condition of the dark 10h of light 14h/, obtain flower training seedling, treat that bloom control is transferred in root media and carried out squamous subculture after growing to 2cm, prescription of rooting medium is: a large amount of+Ms trace+Ms organic matter+sucrose 3%+ agar 0.8% of 1/2Ms and the pH value of root media is 5.8, grow to until bloom control and to transplant after 8cm to Turnover Box strong sprout, after 10 ~ 15 days, be transplanted into land for growing field crops.
(5) bloom control individual plant sowing obtains flower training family, and further economical character, false smut resistance screening obtain the Keng rice lines of improvement:
Field planting bloom control, the japonica rice training family that individual plant sowing obtains the improvement of false smut resistance is some, do not suit training family further field seeding, and to treat that improvement japonica rice variety/be parent is for contrast, screening obtains the Keng rice lines that the good and false smut resistance of agronomic shape obtains improveing.
Embodiment
(1) S0735 and Shan excellent 45 is screened respectively as Japonica two line hybrid rice parent and the anti-source of long-grained nonglutinous rice false smut:
Breeding practice for many years finds, breeding trunk strain S0735 has that output is high, the feature of good grain quality, simultaneously, the plant height of S0735 measures for 2 years on average only has 98.2cm, the time of infertility only has 153 days, sense false smut, and 2011 positive gets S0735 flower pesticide season and be inoculated in and arrange Xian medium and carry out Fiber differentiation, experiment finds that the inductivity of S0735 callus is the same up to 53%(anther culture operating procedure), be applicable to very much the requirement of the Japonica two line hybrid rice parent of this technology path.The Indica Hybrid early rice of Shan You45Shi Hubei Province Xianning institute of agricultural sciences Zhenshan 97a and the seed selection of early indica restorer line 45-10 combo, there is the features such as output is high, rice is of fine quality, the ripe phase is short, disease resistance is strong, how to, multiple false smut fungus strain apparent altitude resistances of false smut lesion, are one of important anti-sources of anti-false smut breeding.
(2) S0735 is carried out to the qualification of Raw toxin patience concentration:
The qualification of Raw toxin endurance concentration was carried out to the S0735 chosen in 2011,0 is added in differential medium, 5%, the Extracted toxin liquid of 10%, 15%, 20%, 25%, 30%, preparation differential medium, differential medium formula is: MS minimal medium+2mg/L6-BA+0.2mg/LNAA+ gradient concentration Extracted toxin liquid and pH value 6.0, then shift into differential medium by the anther callus of Fiber differentiation in step (1), statistics phenylacetic acid, result is as following table one:
Therefore, the Extracted toxin liquid concentration of 24% is decided to be three friendship F1 crossbreed anther culture Raw toxin Selective Pressure by us.
(3) crossbreeding obtains three and hands over F1 crossbreed:
Shans in 2011 excellent 45 and Wide compability japonica rice 02428 of short stem hybridizes builds bridge, and gathers in the crops Hybrid F1, and Hainan in 2012 is chosen F1 flower pesticide and hybridized with S0735 and obtain crossbreed three and hand over F1.
(4) F1 crossbreed is handed over to carry out row Xian medium anther culture to three:
Positive season in 2012, land for growing field crops general planting three handed over F1 crossbreed, choose healthy and strong main Honoka medicine to be inoculated in row's Xian medium and to carry out flower pesticide Fiber differentiation and obtain anther callus, row Xian culture medium prescription is: N6 macroelement+N6 trace element+N6 organic matter+2mg/L2,4-D+0.5mg/LNAA+0.5mg/L protein hydrolysate+5% sucrose+0.8% plant gel, pH value 5.8; And the anther callus selecting diameter 0.4 ~ 0.8cm is inoculated in the differential medium that with the addition of 24% Extracted toxin liquid, differential medium formula is: MS mineral salt+MS organic matter+sucrose 3%+ agar 0.7%+KT2.0mg/L+NAA0.5mg/L+ protein hydrolysate 1g/L+24% Extracted toxin liquid, pH value 6.0, cultivate under the condition of temperature 27 ~ 29 DEG C, the dark 10h of light 14h/, obtain flower training seedling; Treat that bloom control is transferred in root media and carried out squamous subculture after growing to 2cm, prescription of rooting medium is: a large amount of+Ms trace+Ms organic matter+sucrose 3%+ agar 0.8% of 1/2Ms and the pH value of root media is 5.8, growing to until bloom control to transplant to Turnover Box strong sprout after 8cm, within 2013, be transplanted into greenhouse, obtain 127 flower training seedling.
(5) flower training family field planting, economical character, false smut resistance screening obtain 3 excellent Keng rice lines of false smut resistance improvement:
Be transplanted to the bloom control individual plant sowing in greenhouse, obtain 127 flower training familys, positive season in 2014 further field seeding 127 flower training familys take S0735 as contrast, therefrom screen the Keng rice lines that the good and false smut resistance of acquisition 3 agronomic shapes obtains improvement.
Breeding method of the present invention, compared with conventional breeding methods, by special for long-grained nonglutinous rice false smut resistance and import japonica rice efficiently, can greatly reduce screening operation amount, within about 3 years, can complete breeding, significantly shorten conventional anti-false smut breeding cycle; Compared with molecular engineering breeding, there is again the advantage that operation threshold is relatively low, spend less and breeding cycle to shorten, there is larger Breeding Application and be worth.
Above embodiment is only and technological thought of the present invention is described, can not limit protection scope of the present invention with this, every technological thought proposed according to the present invention, and any change that technical scheme basis is done, all falls within scope; The technology that the present invention does not relate to all is realized by prior art.
Claims (10)
1. long-grained nonglutinous rice false smut resistance efficiently imports a breeding method for japonica rice, it is characterized in that: the step of this breeding method is as follows:
(1) choose and wait to improve japonica rice variety/be and the anti-anti-source of false smut long-grained nonglutinous rice: choose the good japonica rice variety of economical character/be and to the japonica rice variety chosen/be the Fiber differentiation carrying out arranging Xian medium, final selected induction of anther callus rate higher than 25% japonica rice variety/be as treating improvement japonica rice variety/be, choose the good and rice variety containing false smut high resistance gene of economical character/be as the anti-anti-source of false smut long-grained nonglutinous rice;
(2) improvement japonica rice variety/be the qualification carrying out Raw toxin patience concentration is treated to what choose: the Extracted toxin liquid adding certain gradient concentration respectively in differential medium, differentiation cultivation is carried out to the anther callus waiting to improve japonica rice variety/be selected in step (1), and selects the Extracted toxin liquid of suitable screening pressure level concentration as japonica rice anther culture Raw toxin Selective Pressure;
(3) crossbreeding obtains three and hands over F1 crossbreed: the anti-anti-source of false smut long-grained nonglutinous rice of screen field planting and Wide compability material Xian round-grained rice are built bridge and obtained Hybrid F1, the flower pesticide choosing Hybrid F1 again with wait to improve japonica rice variety/be and hybridize and obtain crossbreed three and hand over F1;
(4) F1 crossbreed is handed over to carry out row Xian medium anther culture to three: three hand over the positive season sowing of F1 crossbreed, choose healthy and strong main Honoka medicine and carry out row Xian medium flower pesticide Fiber differentiation, obtain anther callus, and the anther callus selecting diameter 0.4 ~ 0.8cm is inoculated in the differential medium that with the addition of Selective Pressure concentration Raw toxin, temperature 27 ~ 29 DEG C, cultivate under the condition of the dark 10h of light 14h/, obtain flower training seedling, treat that bloom control is transferred in root media and carried out squamous subculture after growing to 2cm, growing to until bloom control to transplant to Turnover Box strong sprout after 8cm, land for growing field crops is transplanted into after 10 ~ 15 days,
(5) bloom control individual plant sowing obtains flower training family, further economical character, false smut resistance screening obtain the Keng rice lines of improvement: field planting bloom control, the japonica rice training family that individual plant sowing obtains the improvement of false smut resistance is some, the further field seeding of training family of not suiting, to treat that improvement japonica rice variety/be parent is for contrast, screening obtains the Keng rice lines that the good and false smut resistance of agronomic shape obtains improveing.
2. long-grained nonglutinous rice false smut resistance according to claim 1 efficiently imports the breeding method of japonica rice, it is characterized in that: the japonica rice variety that the economical character in described step (1) is good/be refers to that output is high, good grain quality, false smut resistance need to be improved and plant height should not too high, breeding time is precocious or in ripe japonica rice variety/be; The anti-source of anti-long-grained nonglutinous rice refers to that output is high, the rice variety of good grain quality, high resistance false smut/be.
3. long-grained nonglutinous rice false smut resistance according to claim 1 efficiently imports the breeding method of japonica rice, it is characterized in that: the process that the japonica rice variety that the economical character chosen in described step (1) is good/be carries out the Fiber differentiation arranging Xian medium is as follows:
(1.1) to draw materials and pretreatment: by japonica rice variety good for the economical character chosen/be land for growing field crops conventional cultivation, get pulvinus apart from be 5 ~ 7cm, fringe look be pistac, flower pesticide is faint yellow and length be about clever shell 1/3 ~ 1/2, the main fringe of stalwartness that do not split of booting is for for trying material, retain 2 ~ 3 blades, behind 75% alcohol wipe surface, wrap up and overcoat plastic sack with wet towel, Cold pretreatment 3 days at putting 4 DEG C;
(1.2) inoculate: after pretreatment, rice bud containing main fringe is cut, then this rice bud is soaked in the alcohol of 75% and processes 3 ~ 4min, then this rice bud is shifted and peel off main fringe in superclean bench, strip uniform ramuscule stalk sized by small ear further, the following rear concentration of gauze parcel ramuscule stalk of sterilizing is the mercuric chloride sterilizing 10min of 0.1%, rinsed with sterile water 3 ~ 4 times, open gauze to dry, finally cut off flower pesticide top with the scissors of sterilizing, tremble medicine method inoculation flower pesticide in row Xian medium;
(1.3) Fiber differentiation: dark surrounds evoked callus row Xian medium being placed in 24 ~ 26 DEG C, white-flaxen unformed cell mass is there is at the edge of flower pesticide after 30 ~ 40 days, and add up induction of anther callus rate afterwards flower pesticide inoculated and cultured 50 days, selected induction of anther callus rate higher than the good japonica rice variety of the economical character of 25%/be as treating improvement japonica rice variety/be.
4. long-grained nonglutinous rice false smut resistance according to claim 3 efficiently imports the breeding method of japonica rice, it is characterized in that: the rice bud in described step (1.2) refers in the region of rice bud top pillow distance for internode top 0.5cm under 7cm to fringe the part of cutting material and obtaining, and the upper flower pesticide quantity of the ramuscule stripped stalk is 15 ~ 25.
5. long-grained nonglutinous rice false smut resistance according to claim 3 efficiently imports the breeding method of japonica rice, it is characterized in that: the row Xian culture medium prescription in described step (1.2) is: N6 macroelement+N6 trace element+N6 organic matter+2mg/L2,4-D+0.5mg/LNAA+0.5mg/L protein hydrolysate+5% sucrose+0.8% plant gel, the pH value of row Xian medium is 5.8.
6. the long-grained nonglutinous rice false smut resistance according to claim 1 or 3 efficiently imports the breeding method of japonica rice, it is characterized in that: the Raw toxin patience concentration qualification process in described step (2) is as follows:
(2.1) preparation of Raw toxin: mixing rice false smut is inoculated in plane expand breeding culture medium expands numerous, be transferred to after growing mycelium and produce spore expand in PDA liquid nutrient medium numerous, 28 DEG C of lucifuge shaken cultivation use sterile gauze elimination bacterium after 20 days, culturing filtrate is after the centrifugal 15min of 3000rpm, get supernatant and make crude extract at 121 DEG C of high pressure moist heat sterilization 20min, institute's bacteria-free filtrate that obtains and rice false smut Raw toxin;
(2.2) treat that improvement japonica rice variety/be carries out the qualification of Raw toxin endurance concentration to what choose: the Extracted toxin liquid adding certain gradient concentration respectively in differential medium, then the anther callus waiting to improve japonica rice variety/be Fiber differentiation selected in step (1) is shifted into differential medium, Fiber differentiation added up phenylacetic acid after 30 ~ 40 days, and selected the Extracted toxin liquid of suitable screening pressure level concentration to hand over F1 crossbreed anther culture Raw toxin Selective Pressure concentration as three.
7. long-grained nonglutinous rice false smut resistance according to claim 6 efficiently imports the breeding method of japonica rice, it is characterized in that: the suitable screening pressure level concentration in described step (2.2) refers to the selected Raw toxin concentration of anther callus when being 0.8% ~ 1.2% containing the differentiation rate in the differential medium of Raw toxin until improvement japonica rice variety/be Fiber differentiation.
8. long-grained nonglutinous rice false smut resistance according to claim 1 efficiently imports the breeding method of japonica rice, it is characterized in that: the plane in described step (2.1) expands breeding culture medium formula and is: yeast extract 2.5g/L+ soluble starch 10g/L+ agar powder 16g/L, and pH value is 6.0; PDA liquid culture based formulas is potato 200g/L+ glucose 20g/L+ agar 17g/L, and pH value is 6.5; Differential medium formula in described step (2.2) is: MS minimal medium+2mg/L6-BA+0.2mg/LNAA+ gradient concentration Extracted toxin liquid, the pH value of differential medium is 6.0.
9. long-grained nonglutinous rice false smut resistance according to claim 1 efficiently imports the breeding method of japonica rice, it is characterized in that: three in described step (4) hand over the induction of anther callus rate of F1 crossbreed lower than 3% time, F1 crossbreed and japonica rice parent need be handed over to backcross by three and obtain BC1F1 crossbreed, the positive season sowing of BC1F1 crossbreed, choose healthy and strong main fringe and re-start row Xian medium flower pesticide Fiber differentiation, until obtain the anther callus that inductivity is not less than 3%, the anther callus selecting diameter 0.4 ~ 0.8cm is inoculated in the differential medium adding Raw toxin Selective Pressure, temperature 27 ~ 29 DEG C, cultivate under the condition of the dark 10h of light 14h/, obtain flower training seedling, treat that bloom control is transferred in root media and carried out squamous subculture after growing to 2cm, growing to until bloom control to transplant to Turnover Box strong sprout after 8cm, land for growing field crops is transplanted into after 10 ~ 15 days.
10. long-grained nonglutinous rice false smut resistance according to claim 1 efficiently imports the breeding method of japonica rice, it is characterized in that: the row Xian culture medium prescription in described step (4) is: N6 macroelement+N6 trace element+N6 organic matter+2mg/L2,4-D+0.5mg/LNAA+0.5mg/L protein hydrolysate+5% sucrose+0.8% plant gel, the pH value of row Xian medium is 5.8; Differential medium formula is: the Extracted toxin liquid of MS mineral salt+MS organic matter+suitable screening pressure level concentration of sucrose 3%+ agar 0.7%+KT2.0mg/L+NAA0.5mg/L+ protein hydrolysate 1g/L+, and the pH value of differential medium is 5.8; Prescription of rooting medium is: a large amount of+Ms trace+Ms organic matter+sucrose 3%+ agar 0.8% of 1/2Ms, the pH value of root media is 5.8.
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| CN106106127A (en) * | 2016-06-29 | 2016-11-16 | 无锡南理工科技发展有限公司 | A kind of Folium Sesami pinta resistance Japonica two line hybrid rice method |
| CN106086196B (en) * | 2016-06-29 | 2019-09-20 | 青岛欧易生物科技有限公司 | A kind of rice green smut Resistance detecting method |
| CN114041419A (en) * | 2021-12-15 | 2022-02-15 | 黑龙江省农业科学院生物技术研究所 | Breeding method of new high-quality rice variety with false smut resistance |
| CN114041419B (en) * | 2021-12-15 | 2023-02-28 | 黑龙江省农业科学院生物技术研究所 | Breeding method of new high-quality rice variety with false smut resistance |
| CN115530071A (en) * | 2022-10-13 | 2022-12-30 | 四川省内江市农业科学院 | Breeding method of false smut-resistant rice variety |
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